A PP1-binding motif present in BRCA1 plays a role in its DNA repair function.
Protein phosphatase 1alpha (PP1alpha) regulates phosphorylation of BRCA1, which contains a PP1-binding motif (898)KVTF(901). Mutation of this motif greatly reduces the interaction between BRCA1 and PP1alpha. Here we show that mutation of the PP1-binding motif abolishes the ability of BRCA1 to enhance survival of Brca1-deficient mouse mammary tumor cells after DNA damage. The Rad51 focus formation and comet assays revealed that the DNA repair function of BRCA1 was impaired when the PP1-binding motif was mutated. Analysis of subnuclear localization of GFP-tagged BRCA1 demonstrated that mutation of the PP1-binding motif affected BRCA1 redistribution in response to DNA damage. BRCA1 is required for the formation of Rad51 subnuclear foci after DNA damage. Mutation of the PP1-binding motif in BRCA1 also affected recruitment of Rad51 to sites of DNA damage. Consistent with these findings, knockdown of PP1alpha in BRCA1-proficient cells by small interfering RNA also significantly reduced Rad51 focus formation induced by DNA damage. Further analysis indicated that mutation of the PP1-binding motif compromised BRCA1 activities in homologous recombination. Altogether, our data implicate that interaction with PP1alpha is important for BRCA1 function in DNA repair.
Yu
Young-Mi
YM
Department of Obstetrics, Gynecology and Reproductive Sciences, School of Medicine, University of Pittsburgh, Magee-Womens Research Institute, Pittsburgh, PA15213, USA.
Pace
Serena M
SM
Allen
Susan R
SR
Deng
Chu-Xia
CX
Hsu
Lih-Ching
LC
D020816:Amino Acid Motifs / Q000502:physiology; D000818:Animals; D019313:BRCA1 Protein / Q000032:analysis / Q000737:chemistry* / Q000502:physiology; D001665:Binding Sites; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D004249:DNA Damage; D004260:DNA Repair / Q000502:physiology*; D049452:Green Fluorescent Proteins / Q000032:analysis; D006801:Humans; D008325:Mammary Neoplasms, Experimental / Q000235:genetics / Q000473:pathology; D051379:Mice; D009154:Mutation; D010766:Phosphorylation; D054645:Protein Phosphatase 1 / Q000378:metabolism*; D051135:Rad51 Recombinase / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000032:analysis; D011995:Recombination, Genetic
D019313:BRCA1 Protein; C492913:BRCA1 protein, human; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D051135:Rad51 Recombinase; D054645:Protein Phosphatase 1